Chronic lymphocytic leukemia (CLL) is a malignancy of bone marrow cells (lymphocytes) that is found mostly in older people. It affects about 8,200 Americans each year, representing about 22-30% of all leukemia cases according to the National Cancer Institute. Many patients with CLL live for a long time without treatment and survive many years after diagnosis. Others, however, have disease that rapidly progresses and leads to death within a few years.
The clinical problem has been how to distinguish between these two groups of patients in order to treat each group appropriately. Since the late '90 s it has been known that the presence or absence of somatic mutations in the immunoglobulin heavy chain variable (IgVH) regions of CLL cells provides a reliable distinguishing characteristic between these two patient populations. Patients whose CLL cells express un-mutated IgVH regions usually have early progression of their disease, whereas patients with mutated IgVH regions usually have later progression.
However, one major shortcoming of using IgVH mutations as a biomarker for CLL is that most clinical laboratories are not capable of detecting IgVH mutations. Even if readily available, the technology is expensive to implement and time consuming, making it impractical for widespread screening of all CLL patients. Thus, scientists have been searching for a reliable surrogate marker for IgVH mutation.
Rosenwald found such a marker. In 2001 his group reported that Zeta-associated protein of 70 kDa (ZAP-70), a cytoplasmic tyrosine kinase essential for T-cell-receptor signal transduction, is preferentially expressed in CLL B-cells whose immunoglobulin genes have not undergone somatic hypermutation, while a second CLL subtype with mutated immunoglobulin genes most often lacks ZAP-70 expression. Rosenwald (2001); US2003203416.
Patients with less ZAP-70 in their B-cells were more likely to have mutations, and lived anywhere from 15 to 34 years after their diagnosis, with an average survival of 24.4 years. On the other hand, CLL patients whose cells contained significant ZAP-70 were less likely to have mutations and lived from 7 to 11.5 years, with an average of 9.3 years. Recently, it was shown that ZAP-70 expression in T-cells correlates with B-CLL ZAP-70 and bears the same correlation with time to clinical progression of the disease (Herishanu 2005).
The use of ZAP-70 as a marker for IgVH mutation was initially very promising, however, subsequent studies have cast doubt on its prognostic value. Researchers have criticized the current ZAP-70 assays, as follows:                However, for both CD38 and ZAP-70, subsequent studies have yielded controversial results with regard to their validity as a surrogate marker for VH and prognostic indicator. The facts that (1) divergent results have been obtained in different laboratories (CD38 and ZAP-70), (2) the expression level may change over time (CD38), (3) a careful separation of T-cells is necessary (ZAP-70), (4) different cut-off values to distinguish “positive” from “negative” cases were defined (CD38 and ZAP-70), and (5) approximately 10%-30% of cases show discordant status for CD38 or ZAP-70 as compared to VH in all series described, indicate that these markers may not be as reliable as initially thought for routine diagnostics. Byrd (2004) (cites omitted) (emphasis added).        
The National Cancer Institute has also noted that the test is not standardized: “This test, which is commercially available, but has not undergone national standardization, has been proposed as a surrogate for the mutational status.” Our own lab has noted that ZAP-70 is present only in low copy number against a large background of noise, and these facts have hindered the realization of the true prognostic value of the ZAP-70 assay. Thus, it is apparent that the use of the ZAP-70 assay as a prognostic tool needs considerable improvement. What is needed in the art is a standardized assay with increased reliability, sensitivity and greater quantitative precision.